Project Summary/Abstract. IgG4-related diseases (IgG4-RDs) are a family of related autoinflammatory diseases characterized by fibrotic lesions comprised of CD4+ T cells and IgG4+ plasma cells, and markedly elevated oligoclonal IgG4. However, whether IgG4 antibodies contribute to IgG4-RD pathology remains unclear. Despite the tremendous clinical success of immunotheraputics, little is known regarding IgG4 biology relative to IgG1. Our long-term goals are to understand the role and regulation of glycosylation to antibody biology, to ultimately modulate antibody effector functions in vitro and in vivo. The overall objective of this application is to comprehensively dissect cytotoxic activity of sialylated IgG4. Our central hypothesis is autoantigen-specific IgG4 in IgG4-RD is sialylated, and contributes to the pathology Of IgG4-RD. Our approach combines characterizing IgG from the sera of IgG4-RD and healthy patients, while examining precisely glycoengineered IgG4 in receptor binding and effector function assays. Our hypothesis is informed by preliminary data shown here in the Approach subsection of the Research Strategy section. The rationale that underlies the proposed research is understanding how IgG4 mediate cytotoxic effector function will enable new insights into IgG biology, and may lead to development of innovative antibody-based therapies. We will test our central hypothesis and, thereby, attain the objective of this application by pursuing the following three specific aims using a combination of biophysical experiments, and in vitro and in vivo functional assays. 1) Define the glycosylation and Fc?Rs-binding profiles of total and autoantigen-specific IgG in IgG4-RD. Hypothesis: Sialylation enables Fc?R binding by IgG4. 2) Examine the effector functions of sialylated IgG4 in vitro. Hypothesis: Sialylated IgG4 mediates effector cell-specific pro-inflammatory effector functions. 3) Determine the in vivo effector functions of sialylated IgG4. Hypothesis: Sialylated IgG4 exerts pro- inflammatory effector functions in vivo. This proposal is expected to have broad clinical implications, ranging from diseases where elevated IgG4 titers are associated in the pathology or resolution, as well as to design of immunotherapeutics, and represents a substantive departure from the status quo by underscoring the cytotoxic capacity of IgG4.